The investigations presented in this proposal aim to elucidate the genetic organization, gene expression, and cellular transformation of human papillomaviruses (HPV) by cell culture studies. This elucidation will be achieved by detailed genetic mapping of viral transcripts in cells morphologically transformed by HPV and by investigations of HPV DNA-derived eukaryotic cloning vectors for expression of viral and inserted genetic elements. The HPV-5 type of papillomavirus isolated from epidermodysplasia verruciformis patients with naturally-occurring warts and squamous cell carcinomas will be preferentially used for these studies because of the malignant potential of the virus and the availability of naturally-occurring mutants. Preliminary studies have established the morphological transformation and episomal persistence of transfecting papillomavirus genomes (including HPV-5) in C127 and NIH 3T3 cells. The expression of HPV-5-specific sequences in the latter cell types as well as in novel culture systems potentially permissive for HPV (due to the association of permissive papillomavirus expression with epidermal cell differentiation), granulation tissue cultures and teratocarcinoma cell lines, will be analyzed by RNA mapping procedures and by characterization of infected or transfected cells. HPV-5-specific sequences will also be examined in appropriate cell types transfected with constructed HPV-5 vectors containing inserted genetic elements including the SV40 viral promoter, the herpesvirus thymidine kinase gene, and nonselectable human gene. The outlined studies will provide significant molecular and medically useful information concerning HPV expression.